RNA-binding protein 14 promotes phase separation to sustain prostate specific antigen expression under androgen deprivation in human prostate cancer.

Castration-resistant prostate cancer (CRPC) is a big challenge in managing prostate cancer patients. The androgen receptor (AR) pathway is a major driver even in CRPC under androgen deprivation. The mechanism in maintaining of the AR pathway under androgen deprivation remains elusive. The recent discovery of biomolecular condensate, a membrane-less intracellular construct formed by liquid-liquid phase separation (LLPS), that facilitate molecular assembly, encouraged the re-screening of our previous microarray data list. We selected Rbm14 as a target molecule for further analysis because it works as a coactivator of nuclear receptors as well as it facilitates formation of biomolecular condensates via its intrinsically disordered region. GFP-tagged Rbm14 transfected into HEK293T cells formed droplet-like puncta, which diminished following treatment with 1,6-hexanediol. Droplet-like structures were also observed in immunofluorescence for endogenous RBM14 of PC-3 and DU145 cells. Luciferase assay revealed that Rbm14 enhanced androgen-responsive element (ARE)-mediated reporter activity in all conditions with or without testosterone and AR. Co-immunoprecipitation confirmed the Rbm14-AR interaction. Long non-coding RNAs, including NEAT1, SRA1, and HOTAIR, were also interacted with Rbm14. Small interfering RNAs of NEAT1 reduced ARE-mediated reporter activity, while transfection of SRA1 and HOTAIR enhance the reporter activity. Treatment with 1,6-hexanediol as well as transfection with a dominant-negative splice variant of Rbm14 reduced expression of prostate specific antigen (PSA), a prototype of androgen-regulated gene, in LNCaP, PC-3, and DU145 cells under androgen deprivation. Immunohistochemically, RBM14 expression was substantially upregulated in prostate cancer tissues after androgen deprivation therapy than in untreated tumors. In conclusion, RBM14 is a novel factor involved in maintenance of PSA expression via phase separation under androgen deprivation in prostate cancer.

[Incidentally Detected Right Atrial Blood Cyst].

A 48-year-old man underwent computed tomography for the examination of lower back pain, which incidentally detected a cardiac tumor in the right atrium. On echocardiography, the tumor was identified as a 30 mm round mass with a thin wall and iso- and hyper-echogenic contents that originated from the atrial septum. The tumor was successfully removed under cardiopulmonary bypass, and the patient was discharged in good health. The cyst was filled with old blood, and focal calcification was observed. Pathological examination revealed that the cystic wall was composed of thin-layered fibrous tissue lined with endothelial cells. Regarding a treatment, it is reported that early surgical removal is preferable to avoid embolic complications, however it is controversial. Furthermore, it needs to discuss about the difference between fetal/neonatal and adult cases.

Protective effect of teriparatide against vancomycin-induced cytotoxicity in osteoblasts.

BACKGROUND: Intrawound vancomycin powder is effective in preventing surgical site infection after spine surgery. In a previous study, vancomycin-induced cytotoxicity in osteoblasts was investigated in vitro, and vitamin D3 was verified to be a candidate drug aiding recovery from vancomycin-induced cytotoxicity. The treatment practices involving osteogenesis-promoting drugs vary widely. Teriparatide, an anabolic agent, highly promotes bone formation by inducing osteoblast activation, increasing bone formation and mineral density, and preventing vertebral fractures. Hence, teriparatide may be administered in combination with vancomycin. METHODS: MC3T3-E1 cells were cultured in minimum essential medium supplemented with 10% fetal bovine serum at 37 °C in a humidified incubator containing 5% CO2. The experimental concentrations of vancomycin (2500, 5000, and 7500 µg/mL) were determined based on previous reports and our preliminary experiments. Teriparatide (100 ng/mL) was administered concomitantly to prevent cytotoxicity in osteoblasts, using pulsed vancomycin for 24 h (measured at 1, 3, and 7 days). Cell numbers and morphological changes in cells treated with vancomycin or vancomycin plus 100 ng/mL teriparatide were measured. Osteoblast differentiation was assessed using alkaline phosphatase staining, alkaline phosphatase activity, and alizarin red S staining. RESULTS: Teriparatide showed a recovery effect when vancomycin (7500 µg/mL) was administered only for 24 h. Microscopic examination revealed that teriparatide had a protective effect on osteoblasts exposed to 7500 µg/mL vancomycin. Addition of teriparatide led to the recovery of alkaline phosphatase staining and alizarin red staining. CONCLUSION: Vancomycin-induced cytotoxicity in osteoblasts could be inhibited by administering teriparatide concomitantly with vancomycin.

Development of Low-Molecular-Weight Compounds Targeting the Cancer-Associated KLF5 Transcription Factor.

Krüppel-like factor 5 (KLF5) is a potential target for anticancer drugs. However, as an intrinsically disordered protein (IDP) whose tertiary structure cannot be solved, innovative strategies are needed. We focused on its hydrophobic α-helix structure, defined as an induced helical motif (IHM), which is a possible interface for protein-protein interaction. Using mathematical analyses predicting the α-helix's structure and hydrophobicity, a 4-amino-acid site (V-A-I-F) was identified as an IHM. Low-molecular-weight compounds that mimic the main chain conformation of the α-helix with the four side chains of V-A-I-F were synthesized using bicyclic pyrazinooxadiazine-4,7-dione. These compounds selectively suppressed the proliferation and survival of cancer cells but not noncancer cells and decreased the protein but not mRNA levels of KLF5 in addition to reducing proteins of Wnt signaling. The compounds further suppressed transplanted colorectal cancer cells in vivo without side effects. Our approach appears promising for developing drugs against key IDPs.

Solitary adrenal metastasis from salivary duct carcinoma of the parotid gland successfully treated by surgery: A case report.

RATIONALE: Salivary duct carcinoma (SDC) is a rare and highly aggressive cancer with a poor prognosis. SDC demonstrates a potential for invasive growth with early regional and distant metastasis to organs, such as bone, lung, liver, and brain. Because, adrenal gland metastasis from SDC is rare, its treatment options are not well established. Herein, we report a case of SDC metastasis from the parotid gland to the adrenal gland, which was successfully treated by surgery. PATIENT CONCERNS: The patient had an abnormal but painless lump on the right parotid gland. The size of the mass had increased over a period of 3 years. The patient underwent complete removal of the right parotid gland and radical neck dissection followed by adjuvant radiotherapy and chemotherapy. Two years later, a mass was identified in the left adrenal gland by computed tomography. As no local recurrence or metastasis to other organs was observed, the patient underwent adrenalectomy. DIAGNOSES: Metastasis of SDC in the adrenal gland was confirmed by histopathological examination of the adrenalectomized specimen. INTERVENTIONS: After adrenalectomy, the patient was followed-up without adjuvant therapy. OUTCOMES: The patient was well and alive during the 13-month postoperative follow-up period without any complications. LESSONS: Surgical resection of solitary metastatic lesion may show a survival benefit with metastatic SDC.

Myofibroblast phagocytic cutaneous mucinosis: phagocytosis of mucinous substances by myofibroblasts in a distinctive cutaneous mucinosis: A case report.

RATIONALE: Phagocytosis is an important physiological process for eliminating unnecessary substances or dead cells after tissue damage, such as inflammation or infarction. Phagocytosis was previously considered to be mainly performed by professional phagocytotic cells, such as macrophages. In contrast, we previously demonstrated that the phagocytosis of dead cells and unnecessary substances by myofibroblasts is as important as that by professional phagocytotic cells in myocardial infarction. Based on our discovery, we speculated that phagocytosis by myofibroblasts may be a more common pathological phenomenon also in other diseases than previously believed. PATIENT CONCERNS: A 44-year-old male patient with atopic dermatitis developed a cutaneous reddish nodule with an underlying induration on his thigh. INTERVENTIONS: The cutaneous lesion was surgically removed. DIAGNOSES: Histopathological examination demonstrated that the cutaneous lesion had solid infiltration by inflammatory cells, namely, plasma cells, histiocytes, and lymphocytes, in the dermis. The cutaneous lesion included mucinosis in the dermis. Inside the mucinosis, we detected cells with clear areas of mucinous substances. Some of the cells were α-smooth muscle actin-positive myofibroblasts. Electron microscopic images demonstrated that there were collagen bands in the cells with mucinous engulfment. Based on these pieces of evidence, we conclude that these mucinous phagocytotic cells were myofibroblasts, not professional phagocytotic cells, such as macrophages. OUTCOMES: There was no recurrence of the lesion. LESSONS: The clinical appearance of this case resembled that of previously reported solitary cutaneous focal mucinoses. However, our case had distinctive characteristics, such as the phagocytosis of mucinous substances by myofibroblasts, multiple mucinous lesions in a single eruption, and the presence of inflammatory cells, which have not been previously reported. For this distinct cutaneous lesion, a clear dermatological and pathological name has yet to be determined. We propose "myofibroblast phagocytic cutaneous mucinosis" as a candidate name. In addition, our discoveries suggest that phagocytosis by myofibroblasts is not rare but rather is a common pathological phenomenon that has been undetected or unrecognized.

Does Vitamin D3 Prevent the Inhibitory Effect of Vancomycin on Osteoblasts?

BACKGROUND: The utility of vancomycin powder to prevent surgical site infection, mainly in spinal surgery, has been widely examined, and the local administration of vancomycin powder to wounds has been reported to be effective in preventing surgical site infections after spine surgery. However, in vitro studies have shown that high local concentrations of vancomycin may inhibit osteogenesis, although it remains unclear how these high concentrations influence osteoblasts. No candidate drug has been reported to recover cytotoxicity with high concentrations of vancomycin, but we suggest that vitamin D3, which induces osteoblast proliferation, may be administrated concomitantly with vancomycin in these situations. QUESTIONS/PURPOSES: (1) Does a high concentration of vancomycin reduce viable osteoblast numbers in cell culture compared with controls? (2) Does vitamin D3 administration confer a protective effect on osteoblasts when administered with continuous vancomycin? (3) Does vitamin D3 administration confer a protective effect on osteoblasts when administered with pulsed vancomycin (24 hours of administration)? (4) Does vitamin D3 administration confer alkaline phosphatase, mineralization, and gene expression when administered with pulsed vancomycin? METHODS: MC3T3-E1 cells were cultured at 37° C in an α-minimum essential medium supplemented with 10% fetal bovine serum in a humidified incubator containing 5% CO2. The experimental concentrations of vancomycin (2500 µg/mL, 5000 µg/mL, and 7500 µg/mL) were determined based on previous reports and preliminary experiments. We concomitantly administered vitamin D3 (0.01 nM) to prevent cytotoxicity in osteoblasts, using two different treatments: continuous vancomycin administration (measured at 6 hours, 12 hours, 24 hours, and 72 hours) and pulsed vancomycin for 24 hours (measured at 1 days, 3 days, and 7 days). We analyzed cell numbers and morphologic changes in cells treated with vancomycin or vancomycin plus 0.01 nM vitamin D3. Osteoblast differentiation was assessed with alkaline phosphatase staining, alkaline phosphatase activity, and Alizarin red S staining. RESULTS: The number of cells was reduced at 6 hours, 24 hours, 48 hours, and 72 hours in response to continuous vancomycin administration at 7500 µg/mL (at 72 hours, control 14.6 × 10 cells/mL ± 0.260 × 10 cells/mL, vancomycin at 0.917 × 10 cells/mL ± 0.288 × 10 cells/mL, mean difference -13.7 × 10 cells/mL ± 0.388 × 10 cells/mL [95% CI -14.5 to -12.9]; p < 0.001). Vitamin D3 did not have a protective effect when vancomycin was administered continuously at 7500 µg/mL (at 72 hours, vancomycin alone 0.917 × 10 cells/mL ± 0.288 × 10 cells/mL, vancomycin + vitamin D3 1.67 × 10 cells/mL ± 0.310 × 10 cells/mL, mean difference 0.75 × 10 cells/mL ± 0.423 × 10 cells/mL [95% CI -0.127 to 1.63]; p = 0.09).With pulsed administration for only the first 24 hours, the number of cells was reduced at 1 day, 3 days, and 7 days at 7500 µg/mL (at 7 days, control 18.6 × 10 cells/mL ± 1.29 × 10 cells/mL, vancomycin at 3.46 × 10 cells/mL ± 0.292 × 10 cells/mL, mean difference -15.1 × 10 cells/mL ±1.33 × 10 cells/mL [95% CI -17.9 to -12.4]; p < 0.001 for all). However, vitamin D3 had a recovery effect when vancomycin was administered only for 24 hours (cell number with 7500 µg/mL, day 7: vancomycin alone 3.46 × 10 cells/mL ± 0.292 × 10 cells/mL, vancomycin +vitamin D3 10.6 × 10 cells/mL ± 0.900 × 10 cells/mL, mean difference 7.13 × 10 cells/mL ± 0.946 × 10 cells/mL [95% CI 5.16 to 9.09]; p < 0.001).With the addition of vitamin D3, we observed recovery of alkaline phosphatase staining and Alizarin red staining (evidence of calcification) but no difference in the gene expression of Type I collagen (vancomycin alone 0.319 ± 0.0730, vancomycin + vitamin D3 0.511 ± 0.139, mean difference 0.192 ± 0.157 [95% CI -0.483 to 0.867]; p = 0.345), alkaline phosphatase (vancomycin alone 0.532 ± 0.0210, vancomycin + vitamin D3 0.785 ± 0.0590, mean difference 0.253 ± 0.0620 [95% CI -0.0150 to 0.521]; p = 0.0550), and cathelicidin antimicrobial peptide (vancomycin alone 0.885 ± 0.0520, vancomycin + vitamin D3 1.24 ± 0.125, mean difference 0.355 ± 0.135 [95% CI -0.0200 to 0.730]; p = 0.0580). CONCLUSION: We found that 7500 µg/mL of vancomycin is cytotoxic to osteoblasts. Cytotoxicity could be prevented by administering vitamin D3 in combination with vancomycin. CLINICAL RELEVANCE: The high concentrations of vancomycin routinely used clinically raises concerns related to osteoblast cytotoxicity, which may contribute to pseudoarthrosis after spinal surgery. Thus, vitamin D3, which is frequently used to treat osteoporosis, may have efficacy as a concomitantly administered drug by inducing the proliferation of osteoblasts. These results indicate that a combination therapy of vancomycin and vitamin D3 may prevent adverse events such as osteoblast cytotoxicity.

Primary carcinosarcoma of the ureteropelvic junction associated with ureteral duplication: A case report.

RATIONALE: Primary carcinosarcoma of the upper urinary tract is rare. Ureteral duplication is one of the most common urinary tract malformations. Additionally, the association between ureteral duplication and malignancy is unknown. To the best of our knowledge, no cases of malignant tumors diagnosed as carcinosarcoma with ureteral duplication have been reported. We herein report the case of a patient with carcinosarcoma of the ureteropelvic junction associated with incomplete ureteral duplication. PATIENT CONCERNS: A 60-year-old Japanese woman presented with painless gross hematuria. She had a history of total hysterectomy and chemotherapy for endometrioid carcinoma 5 years before. She had no history of occupational chemical exposure. DIAGNOSES: Radiographic imaging revealed right incomplete ureteral duplication, hydronephrosis, and a polypoid tumor in the ureteropelvic junction of the lower moiety of the right kidney. Urine cytology showed a small amount of degenerated atypical epithelial and nonepithelial cells. The transureteral biopsy specimen showed dysplastic urothelial cells and atypical myoid spindle cells. These findings were indefinite for malignancy. INTERVENTIONS: The patient underwent right nephroureterectomy. Pathological examination of the resected tumor showed a biphasic neoplasm composed of carcinomatous and sarcomatous components. The sarcomatous component was immunohistochemically positive for vimentin, desmin, h-caldesmon, and α-SMA and negative for pancytokeratin (AE1/AE3), low molecular weight cytokeratin (CAM 5.2), EMA, E-cadherin, GATA3, uroplakin 2, and p63. Based on these findings, we diagnosed the tumor as carcinosarcoma. OUTCOMES: The postoperative course was uneventful. No additional therapy was administered. The patient has remained alive without recurrence for 21 months since surgery. LESSONS: Carcinosarcoma can arise from ureteral duplication. Although the majority of carcinosarcomas of the upper urinary tract are diagnosed at an advanced stage and have a poor prognosis, some can have a less aggressive course. Further studies are needed to determine the association between ureteral duplication and malignancy.

Pancreatic carcinoma metastasis to a lung carcinoma lesion and pulmonary fibrotic regions, overtaking the stromal microenvironment: A case report.

RATIONALE: Suppression and of cancer metastasis is one of the most important issues in cancer care. Considering the typical clinical course of metastases, cancer cells might prefer certain environments or conditions. However, favorable environments for cancer metastasis have not been clearly identified. We had previously described a case of dual, yet separate, pancreatic and colon cancer, in which the metastatic pancreatic cancer was localized at the invasive portion of the colon cancer. We hypothesized that metastatic pancreatic cancer took over the colon cancer microenvironment. PATIENT CONCERNS: We experienced an another case of double cancer in a 65-year-old man who had lung squamous cell carcinoma and an independent pancreatic adenocarcinoma that metastasized to the liver as well as to the lung cancer lesion and pulmonary fibrotic regions associated with pneumothorax and bronchiolization. INTERVENTIONS: The pneumothorax could not be controlled by conservative treatment. Thus, an emergency surgery with partial resection of the lower lobe of right lung was performed. DIAGNOSES: We found multiple pancreatic cancer metastases in the lung cancer and fibrotic lesions in the surgical specimen. However, we detected no metastasis in normal lung tissues except inside small arteries, although the lung cancer and fibrotic tissue areas were smaller than the normal lung tissue areas in the surgical specimen. OUTCOMES: The patient died 50 days after the surgery. LESSONS: This case may thus provide evidence to strengthen our hypothesis that pancreatic cancer prefers to metastasize to other independent cancer lesions, overtaking the cancer microenvironment constructed by other independent cancers. The lung cancer microenvironment, rich in myofibroblasts and/or cancer-associated fibroblasts, might be suitable for pancreatic carcinoma metastasis. In addition, we propose the hypothesis that compared with normal tissues, noncancerous fibrotic lesions are preferable destinations for cancer metastasis. Furthermore, metastasis of pancreatic carcinoma to lung cancer and fibrotic tissues might be more common, although such cases have not been previously reported.

Percutaneous fixation of avulsion fracture at the plantar lateral base of the first metatarsal using ZipTight Fixation System: A case report.

Isolated avulsion fracture of the peroneus longus tendon insertion at the base of the first metatarsal without injury of the tarsometatarsal joint is very rare. Similar to most avulsion fractures, this type of injury is caused by strong tension exerted by the peroneus longus tendon. The mechanism leading to this lesion and treatment options are not clearly defined. Several surgical techniques have been advocated for this fracture, including excision of an avulsion fragment and open reduction for internal fixation through the medial aspect of the foot or minimal plantar incision. We have described a method of percutaneous fixing of the avulsion fracture at the plantar lateral base of the first metatarsal using the ZipTight Fixation System (Zimmer Biomet Warsaw, Indiana, USA), which offers the advantage of allowing a rigid fixation and minimal invasive surgical technique for a small fragment.

Traveling Time of a translating ribosome along messenger RNA monitored directly on a quartz crystal microbalance.

During translation, the biosynthesis of polypeptides is dynamically regulated. The translation rate along messenger RNA (mRNA), which is dependent on the codon, structure, and sequence, is not always constant. However, methods for measuring the duration required for polypeptide elongation on an mRNA of interest have not been developed. In this work, we used a quartz crystal microbalance (QCM) technique to monitor mRNA translation in an Escherichia coli cell-free translation system in real time. This method permitted us to evaluate the translation of proteins of interest fused upstream of a streptavidin-binding peptide (SBP) fusion protein. The translation of mRNA encoding the SBP fusion protein alone was observed as a mass increase on a streptavidin-modified QCM plate. Addition of the protein of interest resulted in a delay in the mass change corresponding to the traveling time of the ribosome along the coding region of the protein of interest. With this technique, the lengths of coding sequences, codon usages, influences of unique sequences, and various protein-coding sequences were evaluated. The results showed that the traveling time of the translating ribosome depends on the length of the coding region translated but is also affected by the sequence itself. Differences in the time lags for various proteins imply that mRNA coding sequences may regulate gene expression.

Iconic memory-based omnidirectional route panorama navigation.

A route navigation method for a mobile robot with an omnidirectional image sensor is described. The route is memorized from a series of consecutive omnidirectional images of the horizon when the robot moves to its goal. While the robot is navigating to the goal point, input is matched against the memorized spatio-temporal route pattern by using dual active contour models and the exact robot position and orientation is estimated from the converged shape of the active contour models.

Identification and quantification of N(epsilon)-(Hexanoyl)lysine in human urine by liquid chromatography/tandem mass spectrometry.

The identification and quantification of N(epsilon)-(hexanoyl)lysine (N(epsilon)-HEL), which was found from the reactions between lipid hydroperoxide and lysine, from human urine was examined using liquid chromatography/tandem mass spectrometry (LC/MS/MS). The N(epsilon)-HEL in the partially purified urine fraction was identified using LC/MS/MS by several approaches including precursor/product ion scans. The peak found by the multiple-reaction monitoring (MRM) of the collision-induced fragmentation of N(epsilon)-HEL was clearly observed in urine, and the elution position coincided with the synthetic standard N(epsilon)-HEL. The product, estimated N(epsilon)-HEL, was absorbed by a specific antibody to N(epsilon)-HEL. Moreover, N(alpha)-HEL, one of the plausible hexanoyl adducts from the reaction between the N(alpha) moiety of L-lysine and the peroxidized lipid, was hardly detected in urine samples, suggesting that the origin of the N(epsilon)-HEL is the peroxidized lipid-modified proteins but not artificial hexanoylated L-lysine. Using the MRM technique, the amount of urinary N(epsilon)-HEL from the control subjects (observed healthy) was estimated to be 1.58 +/- 0.23 mumol/mol of creatinine. A comparative study of the urinary N(epsilon)-HEL with an oxidative stress marker, 8-oxo-7,8-dihydro-2'-deoxyguanosine, showed a high correlation (r = 0.844) between the two biomarkers. Furthermore, the quantification of N(epsilon)-HEL in the control and diabetic urines revealed that the urinary N(epsilon)-HEL from diabetic subjects (3.21 +/- 0.65 mumol/mol of creatinine) was significantly higher than that from the control subjects.

Nuclear glutathione S-transferase pi prevents apoptosis by reducing the oxidative stress-induced formation of exocyclic DNA products.

We previously found that nuclear glutathione S-transferase pi (GSTpi) accumulates in cancer cells resistant to anticancer drugs, suggesting that it has a role in the acquisition of resistance to anticancer drugs. In the present study, the effect of oxidative stress on the nuclear translocation of GSTpi and its role in the protection of DNA from damage were investigated. In human colonic cancer HCT8 cells, the hydrogen peroxide (H(2)O(2))-induced increase in nuclear condensation, the population of sub-G(1) peak, and the number of TUNEL-positive cells were observed in cells pretreated with edible mushroom lectin, an inhibitor of the nuclear transport of GSTpi. The DNA damage and the formation of lipid peroxide were dependent on the dose of H(2)O(2) and the incubation time. Immunological analysis showed that H(2)O(2) induced the nuclear accumulation of GSTpi but not of glutathione peroxidase. Formation of the 7-(2-oxo-hepyl)-substituted 1,N(2)-etheno-2'-deoxyguanosine adduct by the reaction of 13-hydroperoxyoctadecadienoic acid (13-HPODE) with 2'-deoxyguanosine was inhibited by GSTpi in the presence of glutathione. The conjugation product of 4-oxo-2-nonenal, a lipid aldehyde of 13-HPODE, with GSH in the presence of GSTpi, was identified by LS/MS. These results suggested that nuclear GSTpi prevents H(2)O(2)-induced DNA damage by scavenging the formation of lipid-peroxide-modified DNA.

Formation of N-(hexanoyl)ethanolamine, a novel phosphatidylethanolamine adduct, during the oxidation of erythrocyte membrane and low-density lipoprotein.

The primary amino groups of biomolecules such as aminophospholipids, as well as proteins, are the potential targets of covalent modifications by lipid peroxidation products; however, little attention has been paid to the modification of aminophospholipids such as phosphatidylethanolamine (PE). The purpose of this study is to characterize the formation of a novel modified phospholipid, N-(hexanoyl)phosphatidylethanolamine (HEPE), in the reaction of PE with lipid hydroperoxides using mass spectrometric analyses. Upon reaction of egg PE with 13-hydroperoxyoctadecadienoic acid or other oxidized polyunsaturated fatty acids followed by phospholipase D-mediated hydrolysis, the formation of N-(hexanoyl)ethanolamine (HEEA), a head group of HEPE, was confirmed by isotope dilution liquid chromatography/tandem mass spectrometry. Moreover, increasing HEEA was detected in the hydrolysates of oxidized erythrocyte ghosts and low-density lipoprotein with their increasing lipid peroxidation levels. Collectively, these results suggest that the N-hexanoylated product of phospholipid, HEPE, can be generated during lipid peroxidation and may serve as one mechanism for the covalent modification of aminophospholipids in vivo.

Formation of N-(hexanoyl)ethanolamine, a novel phosphatidylethanolamine adduct, during the oxidation of erythrocyte membrane and low-density lipoprotein.

The primary amino groups of biomolecules such as aminophospholipids, as well as proteins, are the potential targets of covalent modifications by lipid peroxidation products; however, little attention has been paid to the modification of aminophospholipids such as phosphatidylethanolamine (PE). The purpose of this study was to characterize the formation of a novel modified phospholipid, N-(hexanoyl)phosphatidylethanolamine (HEPE), in the reaction of PE with lipid hydroperoxides using mass spectrometric analyses. Upon reaction of egg PE with 13-hydroperoxyoctadecadienoic acid or other oxidized polyunsaturated fatty acids followed by phospholipase D-mediated hydrolysis, the formation of N-(hexanoyl)ethanolamine (HEEA), a head group of HEPE, was confirmed by isotope dilution liquid chromatography/tandem mass spectrometry. Moreover, increasing HEEA was detected in the hydrolysates of oxidized erythrocyte ghosts and low-density lipoprotein with their increasing lipid peroxidation levels. Collectively, these results suggest that the N-hexanoylated product of phospholipid, HEPE, can be generated during lipid peroxidation and may serve as one mechanism for the covalent modification of aminophospholipids in vivo.